Abstract:

Objective To investigate the mechanism of activated protein C to alleviate the mitochondrialenergy metabolism disorder induced by Mycobacterium tuberculosis in human airway epithelial cells by targetingVLA⁃3⁃neutrophil subpopulation. Methods Human bronchial epithelial BEAS⁃2B cells were divided into threegroups：BEAS⁃2B group（normal culture cell line），Mycobacterium tuberculosis induction group（BEAS⁃2B cellculture was infected with 50 bacteria/cell conjugated bacteria），and BEAS⁃2B transfection group（BEAS⁃2B cellswere transfected with lentiviral vector carrying activated protein C）. The binding degree of activated protein C to vla⁃3 was determined by soluble integrin binding. RT qPCR was used to analyze the mRNA expression of IL⁃6，IL⁃8and MCP⁃1. The function of mitochondria was evaluated by fluorescence probe JC⁃1 and mitochondrial membranepotential was measured by biosciences. Cell apoptosis and viability were detected by ELISA and TUNEL. Results VLA⁃3 was more highly expressed than VLA⁃3 and αVβ3（P < 0.05）. The expression of IL⁃6，IL⁃8 and MCP⁃1mRNA in the induction group of Mycobacterium tuberculosis was higher than that in BEAS⁃2B group（P < 0.05）. TheBEAS⁃2B transfection group was more sensitive than IL⁃6，IL⁃8 and MCP in the induction group of Mycobacteriumtuberculosis. 1 mRNA expression was decreased（P < 0.05）. Compared with the Mycobacterium tuberculosis induc⁃tion group，the mitochondrial complex Ⅰ and mitochondrial complex Ⅳ activity were significantly increased in theBEAS⁃B group（P < 0.05），and the mitochondrial complex I and mitochondrial complex Ⅳ activity were decreasedin the Mycobacterium tuberculosis induction group compared with the BEAS⁃B transfection group（P < 0.05）. Themembrane potential of the BEAS⁃2B group was higher than that of the Mycobacterium tuberculosis induced group（P < 0.05），and the membrane potential of the tuberculosis induction group was lower than that of the BEAS⁃2Btransfection group（P < 0.05）. The apoptosis rate of BEAS⁃2B group was lower than that of Mycobacterium tubercu⁃losis induction group（P < 0.05），and the apoptosis rate of Mycobacterium tuberculosis induction group was higherthan that of BEAS⁃2B transfection group（P < 0.05）. The cell viability of the BEAS⁃2B group was higher than thatof the Mycobacterium tuberculosis induction group（P < 0.05），and the cell viability of the tuberculosis inductiongroup was lower than that of the BEAS⁃2B transfection group（P < 0.05）. Conclusion Activated protein C has astronger affinity for integrin VLA3，which can target VLA3 neutrophil subpopulation to reduce the inflammatoryresponse of human airway epithelial cells，improve mitochondrial complex Ⅰ，complex Ⅳ activity，and effectivelyinhibit binding. Bacillus⁃induced decrease in mitochondrial membrane potential.

Key words: activation protein C, integrin vla?3, apoptosis, mitochondrial dysfunction